Hyaluronidase, which is a generic term for enzymes of degrading hyaluronic acid, was at first known as a spreading factor by Duran-Reynals but later, as it was observed to exhibit strong activity to hyaluronic acid (HA), it came to be called hyaluronidase (HAase). This enzyme is classified, according to its action mechanism, into hyaluronate 4-glycanohydrolase (EC 3.2.1.35) distributed in testicles, lysosomes, and bee venoms; hyaluronate 3-glycanohydrolase (EC 3.2.1.36) present in leech; and hyaluronate lyase (EC 4.2.2.1) present in bacteria.
In particular, as hyaluronidases (PH-20) in testicles are bonded to a glycosylphosphatidylinositol (GPI) anchor site on the acrosome part of a sperm, they are essential enzymes of causing fertilization by degrading a thick outer wall layer outside ovum. It was known that β (1-4) linkage between N-acetyl-D-glucosamine and D-glucuronic acid present in hyaluronic acid (HA), chondroitin, and chondroitin sulfates is hydrolyzed by the action of PH-20. General molecular formula of these enzymes is C2455H3775N617O704S21 and their molecular mass is 53870.9 g/mol. In humans, six genes including HYAL1, HYAL2, HYAL3, and PH-20/SPAM1 are associated with the enzymes.
Since 1950's, the broad applications of the hyaluronidases have been comprehensively reviewed. The first application was the subcutaneous injection of parenteral fluids and besides, they have been used for infiltration and block anesthesia to increase the dispersion of steroids and local anesthetics in orthopedic, ophthalmology, plastic surgery, dental, oral surgery, gynecology, and otolaryngology surgery, and used for dispersion of body fluids not to form agglomeration such as hematoma, prevention of peritoneal adhesion, prevention of calculus formation, and treatment of infertility.
The hyaluronidases currently available on the market are those extracted from ovine testicles and then lyophilized. Such unprocessed hyaluronidases are melted in a suitable concentration, filled into vials and then lyophilized for their manufactures. The thus manufactured hyaluronidases contain excess amounts of foreign proteins. Therefore, the hyaluronidases obtained by the prior methods have not only problems with their stability when converted into a solution but also leave huge problems with their application from a practical perspective because their physiological activity is reduced due to a decrease in their stability over time.